Electroporation procedure

Review all necessary materials including antibodies on ice, the Neon transfection system with configured parameters, a pre-warmed 12-well plate with coverslips, and a cell pellet of 8-10×10⁵ cells in PBS.

Carefully remove all PBS from the pelleted cells, ensuring complete removal of the supernatant while retaining the cell pellet.

Dilute the cell pellet in 80 microliters of R-buffer and transfer the entire cell suspension to a fresh Eppendorf tube.

Transfer 10 microliters of the cell suspension to the antibody tube and mix thoroughly to combine the antibody with the cells.

Using the Neon pipette and tip, carefully draw up the antibody-cell mixture while ensuring no air bubbles are present in the tip, as bubbles will cause cell death.

Insert the loaded pipette into the Neon electroporation station and press start to deliver the electrical pulses for transfection.

After electroporation is complete, remove the pipette from the station and dispense the electroporated cells into the pre-warmed medium in the 12-well plate.